Anchors away: the susceptibility and response to infection between native and co-introduced fishes to the alien anchor worm Lernaea cyprinacea Mikayla McCredden A thesis is presented for the Doctor of Philosophy 2016 Murdoch University, Perth, Western Australia Declaration I declare that this thesis is my own account of my research and contains as its main content work which has not previously been submitted for a degree at any tertiary education institution. Mikayla McCredden A portion of Chapter 1has been published as: Lymbery, A.J., Morine, M., Gholipour Kanani, H., Beatty, S.J. and Morgan, D.L. (2014) Co-invaders: the effects of alien parasites on native hosts. International Journal for Parasitology: Parasites and Wildlife 3: 171-177. My contribution to this paper is estimated at 25% and my total contribution to Chapter 2 is estimated at 80%. ii Acknowledgements Many people have been involved in putting this thesis together. First of all I would like to thank my supervisors, Associate Professor Alan Lymbery, Dr David Morgan, Dr Stephen Beatty and Professor Una Ryan, all of whom have have given me valuable help and guidance along the way. I would like to thank all the fish farmers involved and Australia and Pacific Science Foundation for their funding, without them this project would not have been possible. I would also like to thank Rongchange Yang for working with me on my PCR, Stafania Basile for her input into my thesis, Dr Richmond Loh for his help with fish medicine and James Keleher for helping me with my field sampling. I would also like to give a massive thanks to Susan Kueh, for all of her time, effort and fish expertise. Finally, I would like to thank all of my family who have stood by me over the last few years giving me encouragement and support. In particular, I would like to thank my dad, John, for editing my thesis and always being ready to help me in anyway possible, and my husband, Tim, for keeping me going. iii Abstract The introduction of alien fish species and their alien parasites pose one of the most important threats to freshwater fishes throughout the world. The south-west of Australia has a depauperate, although highly endemic freshwater fish fauna. Of the 200 native freshwater fish species in Australia 144 are exclusively confined to freshwater. In the extreme south-west there are only 11 native freshwater fish species and nine of these are endemic to the region. Six of the 11 freshwater fish species have restricted geographic ranges and four are listed as rare or likely to become extinct. In 2008, studies surveying the parasites of freshwater fishes in the South West Coast Drainage Division (SWCDD), reported the introduction of the alien parasite, Lernaea cyprinacea, into freshwater river systems in the region. Lernaea cyprinacea, commonly known as anchor worm, is a parasitic copepod believed to have been brought in to Western Australia with the accidental release of its native host, Carassius auratus (goldfish). It is not native to Australia and, until recently, had only been reported in fish in eastern Australia. First reports of this parasite in the south-west identified it using morphological criteria from four native freshwater fishes: Galaxias occidentalis (western minnow), Nannoperca vittata (western pygmy perch), Bostockia porosa (nightfish) and Tandanus bostocki (freshwater cobbler). The present study aimed to resolve the morphological uncertainty surrounding the taxonomy of the parasite using molecular techniques, specifically PCR and DNA sequencing, and to review the host range and geographic distribution of this invasive species within the south-west of Western Australia. A comparison of the infection success and pathogenicity of L. cyprinacea in a fish species, Nannoperca vittata (pygmy perch), that is endemic to the Southwestern Province Ichthyological, and that to the natural host, Carassius auratus (goldfish), is detailed. Lernaea cyprinacea in south-western Australia had been morphologically identified in previous studies, but had not been identified using molecular tools. Parasite samples examined in this iv study typed as Lernaea cyprinacea at the 28S ribosomal RNA (rRNA) locus. Sequences were identified using Finch TV Version 1.4.0 (Geospiza Research Team 2004-2006) and checked for identity using the nucleotide database, Nucleotide BLAST (http://blast.ncbi.nlm.nih.gov). The parasite appears to have increased its geographic range in the Southwestern Ichthyological Province; in 2008 it was reported in only one river (the Canning River), whereas in the present study it was found in another two rivers (the Murray River and Serpentine River). Lernaea cyprinacea was also found on two more host species, in addition to the four native hosts reported previously; Galaxias occidentalis (western minnow), Nannoperca vittata (western pigmy perch), Bostockia porosa (nightfish), Tandanus bostocki (freshwater cobbler), and now, Pseudogobius olorum (bluespot goby) and Leptatherina wallacei (western hardyhead). In the field, L. cyprinacea was more prevalent on native freshwater fish species than on the natural host C. auratus. The difference in prevalence of L. cyprinacea on native fishes and C. auratus found in field studies may be due to differences in exposure to the parasite or to differences in susceptibility to infection. Laboratory experiments were used to compare the susceptibility to infection of native N. vittata and C. auratus. There was no difference found in the prevalence or intensity of infection on N. vittata or C. auratus, when they were exposed separately. In mixed communities however, a significantly greater proportion of N. vittata were infected compared to C. auratus (0.59 vs. 0.33), and the mean intensity of infection was also greater in N. vittata than in C. auratus (3.0 ± 0.3 vs. 2.2 ± 0.4). Nannoperca vittata and C. auratus also exhibited significant differences in their behavioural reactions to infection, with putative defensive behaviours observed much more frequently in infected C. auratus than in infected N. vittata. Histologically, C. auratus had a greater pathological and inflammatory response to infection than N. vittata. iv Due to the extensive and destructive effects of C. auratus on both native fishes and habitat, the control of C. auratus has become essential. Removal programs have been underway in Western Australia since 2005, however, we know very little about the effects of removal programs for C. auratus on the co-introduced parasite L. cyprinacea. In particular, it has been suggested that if goldfish are a less competent host species than native freshwater fish species, then removal may actually exacerbate the parasite problem by increasing prevalence of infection on native fishes. This study provides no evidence that the removal of goldfish will exacerbate the problem of L. cyprinacea in river systems in south-western Australia. That being said, there is a need to expand this study to examine the comparative infectivity and pathogenicity of L. cyprinacea to other native fish species and, where possible, to monitor parasite infection rates in the field before and after goldfish control programs to ensure that there are no adverse effects from goldfish removal. vi Table of Contents Author’s Declaration .................................................................................................................... ii Acknowledgements ..................................................................................................................... iii Abstract ....................................................................................................................................... iv Table of Contents ....................................................................................................................... vii List of Figures .............................................................................................................................. x List of Tables .............................................................................................................................. xii Chapter 1 General Introduction ............................................................................................. 1 1.1 Threats to freshwater biodiversity ......................................................................... 1 1.2 Southwestern Ichthyological Province .................................................................. 3 1.3 Biodiversity of freshwater fishes in the Southwestern Province ........................... 4 1.4 Invasive freshwater fishes ..................................................................................... 6 1.5 Effects of invasive fish species ............................................................................. 9 1.6 Carassius auratus as an invasive species ............................................................ 10 1.7 Parasites and invasive species ............................................................................. 11 1.7.1 Parasite loss ......................................................................................................... 11 1.7.2 Spill-back and dilution ........................................................................................ 12 1.7.3 Spillover .............................................................................................................. 13 1.8 Virulence of introduced parasites to native hosts................................................ 17 1.9 Control of invasive species and co-invading parasites